It is well known to perform a quantitative or qualitative analysis of an aqueous liquid by contacting that liquid with a combination of reagents capable of yielding a detectable product in proportion to the concentration of the analyte in the liquid. One type of useful assay utilizes enzymatic reactions wherein the analyte, upon contact with the appropriate reagents, reacts with oxygen in the presence of a suitable enzyme to produce hydrogen peroxide in proportion to concentration of the analyte. A detectable product is then produced by the reaction of hydrogen peroxide in proportion to the concentration of the analyte in the test liquid. Peroxidase is generally used in such assays.
In other assays, a peroxidase is reacted in the presence of hydrogen peroxide which has been added to the system to measure the amount of a particular analyte. Analytes such as glucose, triglycerides, uric acid, cholesterol and creatine kinase can be so detected as well as specific binding ligands in specific binding assays wherein the peroxidase is used as a detectable label. Such determinations can be carried out in solution, dry analytical assays or diagnostic test devices. The signals produced in such assays can be a colorimetric, chemiluminescent or fluorescent signal using well known signal generating reagents.
It has been previously demonstrated that certain phenols and anilines can be used to enhance the production of a colorimetric signal using peroxidase as a reagent in signal generation. See, for example, U.S. Pat. No. 4,828,983 (McClune).
There are also several major types of luminescent or luminometric assays which produce an emission of light as a result of the presence of the analyte of interest. These assays are also known as chemiluminescent assays and are described, for example, in U.S. Pat. No. 4,729,950 (Kricka et al) and publications noted therein. Various aromatic amines and phenols, such as p-iodophenol, are considered useful for enhancing the production of light in such assays (see also U.S. Pat. No. 4,598,044 of Kricka et al).
One preferred enhancer of colorimetric assays has been 4'-hydroxyacetanilide, which is described in the McClune patent noted above.
Although 4'-hydroxyacetanilide shows remarkable behavior in many analytical systems, there is a need for further improvement. For example, high concentrations of 4'-hydroxyacetanilide do not produce correspondingly high signal generation. In addition, the stability of the compound needs to be increased.